Oncogenomics and Epigenetics Unit, IRCCS-Regina Elena National Cancer Institute, 00144, Rome, Italy.
Cell and Molecular Biology Department, School of Biology, College of Science, University of Tehran, Tehran, 1417614411, Iran.
J Exp Clin Cancer Res. 2019 Mar 29;38(1):141. doi: 10.1186/s13046-019-1119-x.
Head and neck squamous cell carcinoma (HNSCC) is the sixth leading cancer worldwide. They are typically characterized by a high incidence of local recurrence, which is the most common cause of death in HNSCC patients. TP53 is the most frequently mutated gene in HNSCC and patients carrying TP53 mutations are associated with a higher probability to develop local recurrence. MiRNAs, which are among the mediators of the oncogenic activity of mt-p53 protein, emerge as an appealing tool for screening, diagnosis and prognosis of cancer. We previously identified a signature of 12 miRNAs whose aberrant expression associated with TP53 mutations and was prognostic for HNSCC. Among them miR-96-5p emerges as an oncogenic miRNAs with prognostic significance in HNSCC.
To evaluate the oncogenic role of miR-96-5p in a tumoral context, we performed colony formation, cell migration and cell viability assays in two HNSCC cell lines transfected for miR-96-5p mimic or inhibitor and treated with or without radio/chemo-therapy. In addition, to identify genes positively and negatively correlated to miR-96-5p expression in HNSCC, we analyzed the correlation between gene expression and miR-96-5p level in the subset of TCGA HNSCC tumors carrying missense TP53 mutations by Spearman and Pearson correlation. To finally identify targets of miR-96-5p, we used in silico analysis and the luciferase reporter assay to confirm PTEN as direct target.
Our data showed that overexpression of miR-96-5p led to increased cell migration and radio-resistance, chemotherapy resistance in HNSCC cells. In agreement with these results, among the most statistically significant pathways in which miR-96-5p is involved, are focal Adhesion, extracellular matrix organization and PI3K-Akt-mTOR-signaling pathway. As a direct target of miR-96-5p, we identified PTEN, the main negative regulator of PI3K-Akt signalling pathway activation.
These results highlight a new mechanism of chemo/radio-resistance insurgence in HNSCC cells and support the possibility that miR-96-5p expression could be used as a novel promising biomarker to predict radiotherapy response and local recurrence development in HNSCC patients. In addition, the identification of pathways in which miR-96-5p is involved could contribute to develop new therapeutic strategies to overcome radio-resistance.
头颈部鳞状细胞癌（HNSCC）是全球第六大常见癌症。它们通常以局部复发率高为特征，这是 HNSCC 患者死亡的最常见原因。TP53 是 HNSCC 中突变最频繁的基因，携带 TP53 突变的患者发生局部复发的可能性更高。miRNAs 是 mt-p53 蛋白致癌活性的中介物之一，作为癌症筛查、诊断和预后的有吸引力的工具出现。我们之前确定了一个由 12 个 miRNA 组成的特征，其异常表达与 TP53 突变相关，并对头颈部鳞状细胞癌具有预后意义。其中，miR-96-5p 作为一种具有致癌意义的 miRNA，对头颈部鳞状细胞癌具有预后意义。
为了评估 miR-96-5p 在肿瘤环境中的致癌作用，我们在转染 miR-96-5p 模拟物或抑制剂的两种 HNSCC 细胞系中进行了集落形成、细胞迁移和细胞活力测定，并进行了放射/化疗处理。此外，为了确定与 HNSCC 中 miR-96-5p 表达呈正相关和负相关的基因，我们通过 Spearman 和 Pearson 相关性分析，在携带错义 TP53 突变的 TCGA HNSCC 肿瘤亚集中分析基因表达与 miR-96-5p 水平之间的相关性。最后，我们通过计算机分析和荧光素酶报告基因检测来确认 PTEN 是 miR-96-5p 的直接靶标，从而确定 miR-96-5p 的靶标。
我们的数据表明，miR-96-5p 的过表达导致 HNSCC 细胞的迁移增加和放射抵抗、化疗耐药。与这些结果一致的是，在 miR-96-5p 参与的最具统计学意义的途径中，有焦点粘附、细胞外基质组织和 PI3K-Akt-mTOR 信号通路。作为 miR-96-5p 的直接靶标，我们鉴定出了 PTEN，它是 PI3K-Akt 信号通路激活的主要负调节剂。
这些结果强调了 HNSCC 细胞中化学/放射抵抗新机制的出现，并支持 miR-96-5p 表达可作为预测 HNSCC 患者放疗反应和局部复发发展的新有前途的生物标志物的可能性。此外，确定 miR-96-5p 参与的途径可能有助于开发新的治疗策略来克服放射抵抗。
